Genetic analysis of semen characteristic traits in Norwegian Red bulls at the artificial insemination center.

Compared with cow fertility, genetic analyses of bull fertility are limited and based on relatively few animals. The aim of the present study was to estimate genetic parameters for semen characteristics of Norwegian Red bulls at the artificial insemination (AI) center (Geno AI station, Stange, Norway) and to estimate genetic correlations between some of these traits and andrology traits measured at the performance test station. The data from the AI center consisted of records from 137,919 semen collections from 3,145 bulls with information on semen weight, sperm concentration, motility before and after cryopreservation, motility change during cryopreservation, and number of accepted straws made. Data from the performance test station included 12,522 observations from 3,219 bulls on semen volume, concentration, and motility (%) when fresh and after storing for 24 and 48 h. Genetic parameters were estimated using linear animal repeatability models that included fixed effects of year-month of observation, age of bull, interaction between semen collection number, and interval between collections for all traits and type of diluter for postcryopreservation traits. The random effects included test-day, permanent environmental, and additive genetic effects of the bull. Based on records from the AI center, we found that semen weight, sperm concentration, and number of straws were moderately heritable (0.18-0.20), whereas motility had a lower heritability (0.02-0.08). Heritability of motility (%) was higher after cryopreservation than before. Genetic correlations among the semen characteristics ranged from unfavorable (-0.35) to favorable (0.93), with standard errors ranging from 0.02 to 0.22. Among the most precise genetic correlation estimates, number of straws made from a batch correlated favorably with semen weight (0.62 ± 0.06) and sperm concentration (0.44 ± 0.08), whereas sperm concentration was negatively correlated with weight (-0.33 ± 0.09). The genetic correlation between motility (%) before and after cryopreservation was 0.64 ± 0.14, and motility change during cryopreservation had a strong favorable genetic correlation with motility after cryopreservation (-0.93 ± 0.02). The estimated genetic correlation (standard error) between the traits volume, concentration, and motility when fresh measured at the performance test station and their respective corresponding traits at the AI center were 0.83 (0.05), 0.78 (0.09), and 0.49 (0.31). The final product at the AI center (number of accepted straws) correlated genetically favorably with all semen characteristic traits recorded at the performance test station (ranging from 0.51 to 0.67). Our results show that the andrology testing done at the performance test station is a resource to identify the genetically best bulls for AI production.

Genetic correlations between body weight, daily weight gain, and semen characteristic traits in young Norwegian Red bulls.

The aim of this study was to estimate genetic parameters for body weight (BW) at 150 d (Bw_150d), and 330 d (Bw_330d) of age and average daily weight gain (Dwg), and to estimate genetic correlations between these traits and semen characteristic traits: volume; concentration (Conc); motility in fresh, 24-h, and 48-h samples (Mot0h, Mot24h, Mot48h); and sperm defects. Data were collected at the performance test station of young Norwegian Red bulls from 2002 to 2012, before selection of bulls for artificial insemination. The weight and growth data consisted of observations for 3,209 bulls, and andrology information was available for up to 2,034 of these bulls. Genetic parameters were estimated using linear animal models. Models for BW and growth traits included the group and year the bull left the station and the pen they occupied during weighing (group-year-pen) and parity of their dam as fixed effects. Models for andrology traits had group-year, age in months (11 to 15), and the interaction between ejaculate number and days since previous collection included as fixed effects. Estimated heritability was 0.14 for Bw_150d, 0.26 for Bw_330d, and 0.34 for Dwg; the estimated genetic correlations among these traits were all favorable. Both BW traits correlated favorably with all the semen characteristic traits (0.20 to 0.76), whereas Dwg was favorably correlated with volume, Mot24h, Mot48h, and sperm defects, and unfavorably correlated with Conc (-0.25) and Mot0h (-0.53). Our results indicate that the genetic correlations between weight and growth traits and semen characteristics depend on the age of the bulls. Although most genetic correlations were favorable, selection for higher daily weight gain between 150 and 330 d might explain the slight negative genetic trends observed for semen characteristics in young Norwegian Red bulls.

Genetic analysis of semen characteristic traits in young Norwegian Red bulls.

The aim of this study was to estimate genetic parameters and genetic trends for male fertility in Norwegian Red bulls. We analyzed data on semen characteristics traits collected at the performance test station of young bulls from 1994 to 2016, in an andrology test used to ensure acceptable semen quality before being selected as an artificial insemination bull. Traits included were volume, concentration, and motility (percentage of moving sperm cells) in fresh samples and after storing for 24 and 48 h, and sperm defects. The data consisted of 14,972 ejaculates from 3,927 young (11-15 mo) Norwegian Red bulls. Genetic parameters were estimated using bivariate linear animal models that included age in months, group-year, and collection-group (main effect of the interaction between ejaculate number and interval between collections) as fixed effects, and test-day and additive genetic and permanent environment effect of the bull as random effects. Considerable genetic coefficients of variation were found for concentration and volume, with lower values for motility. Estimated heritabilities ranged from 0.02 and 0.03 (for sperm defects and motility in fresh samples) to 0.14 (volume and concentration measured on a continuous scale). All estimated genetic correlations were favorable, but the genetic correlations between volume and concentration and volume and sperm defects were not significantly different from zero. The genetic correlations between concentration and motility traits ranged from 0.53 to 0.83, and those between volume and the motility traits were between 0.24 and 0.57. All traits showed a slightly unfavorable genetic trend. Our results indicate that selection of bulls with better sperm quality is possible.

Selective activation of intra-muscular compartments within the trapezius muscle in subjects with Subacromial Impingement Syndrome. A case-control study.

Neuromuscular control of the scapular muscles is important in the etiology of shoulder pain. Electromyographical (EMG) biofeedback in healthy people has been shown to support a selective activation of the lower compartment of the trapezius muscle, specifically. The aim of the present paper was to investigate whether patients with Subacromial Impingement Syndrome (SIS) were able to selectively activate the individual compartments within the trapezius muscle, with and without EMG biofeedback to the same extent as healthy controls (No-SIS). Fifteen SIS and 15 No-SIS participated in the study. Sessions with and without visual biofeedback were conducted. Surface EMG was recorded from four compartments of the trapezius muscle. Selective activation was defined as activation above 12% with other muscle parts below 1.5% or activation ratio at or above 95% of the total activation. Without biofeedback significantly fewer SIS subjects than No-SIS achieved selective activation (p=0.02-0.03). The findings of the study show that without biofeedback No-SIS had a superior scapular muscle control. However, when provided with visual EMG feedback the SIS group performed equally well as the No-SIS group. This indicated that individuals with SIS may benefit from biofeedback training to gain control of the neuromuscular function of the scapular muscle.

The use of EMG biofeedback for learning of selective activation of intra-muscular parts within the serratus anterior muscle: a novel approach for rehabilitation of scapular muscle imbalance.

Motor control and learning possibilities of scapular muscles are of clinical interest for restoring scapular muscle balance in patients with neck and shoulder disorders. The aim of the study was to investigate whether selective voluntary activation of intra-muscular parts within the serratus anterior can be learned with electromyographical (EMG) biofeedback, and whether the lower serratus anterior and the lower trapezius muscle comprise the lower scapula rotation force couple by synergistic activation. Nine healthy males practiced selective activation of intra-muscular parts within the serratus anterior with visual EMG biofeedback, while the activity of four parts of the serratus anterior and four parts of the trapezius muscle was recorded. One subject was able to selectively activate both the upper and the lower serratus anterior respectively. Moreover, three subjects managed to selectively activate the lower serratus anterior, and two subjects learned to selectively activate the upper serratus anterior. During selective activation of the lower serratus anterior, the activity of this muscle part was 14.4+/-10.3 times higher than the upper serratus anterior activity (P<0.05). The corresponding ratio for selective upper serratus vs. lower serratus anterior activity was 6.4+/-1.7 (P<0.05). Moreover, selective activation of the lower parts of the serratus anterior evoked 7.7+/-8.5 times higher synergistic activity of the lower trapezius compared with the upper trapezius (P<0.05). The learning of complete selective activation of both the lower and the upper serratus anterior of one subject, and selective activation of either the upper or lower serratus anterior by five subjects designates the promising clinical application of EMG biofeedback for restoring scapular muscle balance. The synergistic activation between the lower serratus anterior and the lower trapezius muscle was observed in only a few subjects, and future studies including more subjects are required before conclusions of a lower scapula rotation couple can be drawn.

Selective activation of neuromuscular compartments within the human trapezius muscle.

Task-dependent differences in relative activity between "functional" subdivisions within human muscles are well documented. Contrary, independent voluntary control of anatomical subdivisions, termed neuromuscular compartments is not observed in human muscles. Therefore, the main aim of this study was to investigate whether subdivisions within the human trapezius can be independently activated by voluntary command using biofeedback guidance. Bipolar electromyographical electrodes were situated on four subdivisions of the trapezius muscle. The threshold for "active" and "rest" for each subdivision was set to >12% and <1.5% of the maximal electromyographical amplitude recorded during a maximal voluntary contraction. After 1h with biofeedback from each of the four trapezius subdivisions, 11 of 15 subjects learned selective activation of at least one of the four anatomical subdivisions of the trapezius muscle. All subjects managed to voluntarily activate the lower subdivisions independently from the upper subdivisions. Half of the subjects succeeded to voluntarily activate both upper subdivisions independently from the two lower subdivisions. These findings show that anatomical subdivisions of the human trapezius muscle can be independently activated by voluntary command, indicating neuromuscular compartmentalization of the trapezius muscle. The independent activation of the upper and lower subdivisions of the trapezius is in accordance with the selective innervation by the fine cranial and main branch of the accessory nerve to the upper and lower subdivisions. These findings provide new insight into motor control characteristics, learning possibilities, and function of the clinically relevant human trapezius muscle.

An analysis of motor unit firing pattern during sustained low force contraction in fatigued muscle.

In the present study motor unit (MU) firing pattern was analysed during long-term static contraction in order to see if fatigue would induce rotation of activity between different MU. Surface as well as intramuscular EMG were obtained from ten subjects during a sustained hand lift for 5 minutes after performance of a 30% MVC fatiguing contraction of the extensor carpi radialis muscle. A newly developed decomposition program constituted a powerful tool to obtain detailed knowledge of long term activity pattern of MU during low force contractions. Although the muscle was highly fatigued the majority of MU showed a continuous firing pattern after recruitment and no clear incidences of rotation were found for any of the subjects. Therefore, long term, low force contractions, as performed during many occupational work tasks, may involve continuous activation of the low threshold MU and this could have mechanical as well as metabolic implications for these muscle fibers.

Doublets in motor unit activity of human forearm muscle during simulated computer work.

Motor unit firing pattern and characteristics were analyzed during voluntary contractions as they occur during computer work. For comparison controlled standard ramp contractions were performed. Highly diverse firing patterns were identified while the MU characteristics were similar. Of special notice is the frequent occurrence of double discharges during computer like work but not during ramp contractions.

Properties of small molecules affecting insulin receptor function.

Small molecules with insulin mimetic effects and oral availability are of interest for potential substitution of insulin injections in the treatment of diabetes. We have searched databases for compounds capable of mimicking one epitope of the insulin molecule known to be involved in binding to the insulin receptor (IR). This approach identifies thymolphthalein, which is an apparent weak agonist that displaces insulin from its receptor, stimulates auto- and substrate phosphorylation of IR, and potentiates lipogenesis in adipocytes in the presence of submaximal concentrations of insulin. The various effects are observed in the 10(-5)-10(-3) M range of ligand concentration and result in partial insulin activity. Furthermore, analogues of the related phenol red and fluorescein molecules fully displace insulin from the IR ectodomain, however, without insulin agonistic effects. The interactions are further characterized by NMR, UV-vis, and fluorescence spectroscopies. It is shown that both fluorescence and UV-vis changes in the ligand spectra induced by IR fragments occur with Kd values similar to those obtained in the displacement assay. Nevertheless, insulin itself cannot completely abolish binding of the small molecules. Determination of the binding stoichiometry reveals multiple binding sites for ligands of which one overlaps with the insulin binding site on the receptor.

Evaluation of intra-muscular EMG signal decomposition algorithms.

We propose and test a tool to evaluate and compare EMG signal decomposition algorithms. A model for the generation of synthetic intra-muscular EMG signals, previously described, has been used to obtain reference decomposition results. In order to evaluate the performance of decomposition algorithms it is necessary to define indexes which give a compact but complete indication about the quality of the decomposition. The indexes given by traditional detection theory are in this paper adapted to the multi-class EMG problem. Moreover, indexes related to model parameters are also introduced. It is possible in this way to compare the sensitivity of an algorithm to different signal features. An example application of the technique is presented by comparing the results obtained from a set of synthetic signals decomposed by expert operators having no information about the signal features using two different algorithms. The technique seems to be appropriate for evaluating decomposition performance and constitutes a useful tool for EMG signal researchers to identify the algorithm most appropriate for their needs.

Motor unit activity during stereotyped finger tasks and computer mouse work.

Motor unit (MU) activity pattern was examined in the right-hand extensor digitorum communis muscle (EDC) during standardised finger movements simulating actual computer mouse tasks. Intramuscular recordings were performed with a quadripolar needle electrode. Nine women performed four lifts of their right-hand index finger, middle finger or both as well as a number of double clicks. Additionally, the subjects performed contra lateral activity with their left-hand fingers and for three subjects recordings were also obtained during an interview with no physical activity. Besides the expected close coupling of MU activity with finger movement, activity was observed in three different situations with no physical requirements. Attention related activity was found before or after performance of the finger movement task, contra lateral activity in right EDC during left-hand finger tasks, and activity during mental activity without any finger movements involved. A relatively large number of doublet occurrences suggest they are a natural part of the activation pattern during performance of the rapid finger movement required to perform an efficient double click on the computer mouse.

Structural effects of protein lipidation as revealed by LysB29-myristoyl, des(B30) insulin.

Intracellular proteins are frequently modified by covalent addition of lipid moieties such as myristate. Although a functional role of protein lipidation is implicated in diverse biological processes, only a few examples exist where the structural basis for the phenomena is known. We employ the insulin molecule as a model to evaluate the detailed structural effects induced by myristoylation. Several lines of investigation are used to characterize the solution properties of Lys(B29)(N(epsilon)-myristoyl) des(B30) insulin. The structure of the polypeptide chains remains essentially unchanged by the modification. However, the flexible positions taken up by the hydrocarbon chain selectively modify key structural properties. In the insulin monomer, the myristoyl moiety binds in the dimer interface and modulates protein-protein recognition events involved in insulin dimer formation and receptor binding. Myristoylation also contributes stability expressed as an 30% increase in the free energy of unfolding of the protein. Addition of two Zn(2+)/hexamer and phenol results in the displacement of the myristoyl moiety from the dimer interface and formation of stable R(6) hexamers similar to those formed by human insulin. However, in its new position on the surface of the hexamer, the fatty acid chain affects the equilibria of the phenol-induced interconversions between the T(6), T(3)R(3), and R(6) allosteric states of the insulin hexamer. We conclude that insulin is an attractive model system for analyzing the diverse structural effects induced by lipidation of a compact globular protein.

The relationship between insulin bioactivity and structure in the NH2-terminal A-chain helix.

Studies of naturally occuring and chemically modified insulins have established that the NH2-terminal helix of the A-chain is important in conferring affinity in insulin-receptor interactions. Nevertheless, the three-dimensional structural basis for these observations has not previously been studied in detail. To correlate structure and function in this region of the molecule, we have used the solution structure of an engineered monomer (GluB1, GluB10, GluB16, GluB27, desB30)-insulin (4E insulin) as a template for design of A-chain mutants associated with enhanced or greatly diminished affinity for the insulin receptor. In the context of 4E insulin, the employed mutants, i.e. ThrA8-->His and ValA3-->Gly, result in species with 143% and 0.1% biological activity, respectively, relative to human insulin. The high-resolution NMR studies reveal two well-defined structures each resembling the template. However, significant structural differences are evident notably in residues A2-A8 and their immediate environment. In comparison with the template structure, the A8His mutation enhances the helical character of residues A2-A8. This structural change leads to additional exposure of a hydrophobic patch mainly consisting of species invariant residues. In contrast, the A3Gly mutation leads to stretching and disruption of the A2-A8 helix and changes both the dimensions and the access to the hydrophobic patch exposed in the more active insulins. We conclude that the mutations induce small, yet decisive structural changes that either mediate or inhibit the subtle conformational adjustments involved in the presentation of this part of the insulin pharmacophore to the receptor.

A structural switch in a mutant insulin exposes key residues for receptor binding.

Despite years of effort to clarify the structural basis of insulin receptor binding no clear consensus has emerged. It is generally believed that insulin receptor binding is accompanied by some degree of conformational change in the carboxy-terminal of the insulin B-chain. In particular, while most substitutions for PheB24 lead to inactive species, glycine or D-amino acids are well tolerated in this position. Here we assess the conformation change by solving the solution structure of the biologically active (GluB16, GlyB24, desB30)-insulin mutant. The structure in aqueous solution at pH 8 reveals a subtle, albeit well-defined rearrangement of the C-terminal decapeptide involving a perturbation of the B20-23 turn, which allows the PheB25 residue to occupy the position normally taken up by PheB24 in native insulin. The new protein surface exposed rationalizes the receptor binding properties of a series of insulin analogs. We suggest that the structural switch is forced by the structure of the underlying core of species invariant residues and that an analogous rearrangement of the C-terminal of the B-chain occurs in native insulin on binding to its receptor.

Solution structure of an engineered insulin monomer at neutral pH.

Insulin circulates in the bloodstream and binds to its specific cell-surface receptor as a 5808 Da monomeric species. However, studies of the monomer structure and dynamics in solution are severely limited by insulin self-association into dimers and higher oligomers. In the present work we use site-directed mutagenesis of the dimer- and hexamer-forming surfaces to yield the first insulin species amenable for structure determination at neutral pH by nuclear magnetic resonance (NMR) spectroscopy. The preferred insulin mutant, i.e., (B1, B10, B16, B27) Glu, des-B30 insulin retains 47% biological potency and remains monomeric at millimolar concentrations in aqueous solution at pH 6.5-7.5 as judged by NMR and near-UV circular dichroism (CD) spectroscopy. From a series of 2D 1H-NMR spectra collected at pH 6.5 and 34 degrees C, the majority of the resonances are assigned to specific residues in the sequence, and nuclear Overhauser enhancement (NOE) cross-peaks are identified. NOE-derived distance restraints in conjunction with torsion restraints based on measured coupling constants, 3JHNH alpha, are used for structure calculations using the hybrid method of distance geometry and simulated annealing. The calculated structures show that the major part of the insulin mutant is structurally well defined with an average root mean square (rms) deviation between the 25 calculated structures and the mean coordinates of 0.66 A for backbone atoms (A2-A19 and B4-B26) and 1.31 A for all backbone atoms. The A-chain consists of two antiparallel helices, A2-A7 and A12-A19, connected by a loop. The B-chain contains a loop region (B1-B8), an alpha-helix (B9-B19), and a type I turn (B20-B23) and terminates as an extended strand (B24-B29). The B1-B4 and B27-B29 regions are disordered in solution. The structure is generally similar to crystal structures and resembles a crystalline T-state more than an R-state in the sense that the B-chain helix is confined to residues B9-B19.

Solution structure of the superactive monomeric des-[Phe(B25)] human insulin mutant: elucidation of the structural basis for the monomerization of des-[Phe(B25)] insulin and the dimerization of native insulin.

The three-dimensional solution structure of des-[Phe(B25)] human insulin has been determined by nuclear magnetic resonance spectroscopy and restrained molecular dynamics calculations. Thirty-five structures were calculated by distance geometry from 581 nuclear Overhauser enhancement-derived distance constraints, ten phi torsional angle restraints, the restraints from 16 helical hydrogen bonds, and three disulfide bridges. The distance geometry structures were optimized using simulated annealing and restrained energy minimization. The average root-mean-square (r.m.s.) deviation for the best 20 refined structures is 1.07 angstroms for the backbone and 1.92 angstroms for all atoms if the less well-defined N and C-terminal residues are excluded. The helical regions are more well defined, with r.m.s. deviations of 0.64 angstroms for the backbone and 1.51 angstroms for all atoms. It is found that the des-[Phe(B25)] insulin is a monomer under the applied conditions (4.6 to 4.7 mM, pH 3.0, 310 K), that the overall secondary and tertiary structures of the monomers in the 2Zn crystal hexamer of native insulin are preserved, and that the conformation-averaged NMR solution structure is close to the structure of molecule 1 in the hexamer. The structure reveals that the lost ability of des-[Phe(B25)] insulin to self-associate is caused by a conformational change of the C-terminal region of the B-chain, which results in an intra-molecular hydrophobic interaction between Pro(B28) and the hydrophobic region Leu(B11)-Leu(B15) of the B-chain alpha-helix. This interaction interferes with the inter-molecular hydrophobic interactions responsible for the dimerization of native insulin, depriving the mutant of the ability to dimerize. Further, the structure displays a series of features that may explain the high potency of the mutant on the basis of the current model for the insulin-receptor interaction. These features are: a change in conformation of the C-terminal region of the B-chain, the absence of strong hydrogen bonds between this region and the rest of the molecule, and a relatively easy accessibility to the Val(A3) residue.